Herpes simplex virus type 2 (HSV-2) is the major cause of genital herpes, one of the most frequent sexually transmitted diseases. With an estimated US seroprevalence rate of 17% among 14-49 years olds, the recognition of genital herpes as a worldwide pandemic, and the increased risk that HSV-2 plays on the HIV-1 epidemic, the development of strategies to prevent HSV infection is at the forefront of herpes research. We have identified 2 groups of subjects who appear to resist infection in the face of frequent HSV-2 exposure from infected sexual partners. The first group of HSVneg subjects we have identified, denoted Exposed Seronegative (ES), include HSV-1 and -2 seronegative subjects in long-standing sexual relationships with HSV-2 infected partners, who have no evidence of HSV infection and/or disease and who lack detectable systemic HSV-specific T cell responses. The second group, denoted Immune Seronegative (IS), includes HSV-1 and -2 seronegative subjects who have current or prior histories of mucosal HSV exposure from infected sexual partners, have no virological or clinical evidence of HSV infection and/or disease, and who possess frequent and persistent systemic HSV-specific CD4 and CD8 T cell responses. While HSV-2+ subjects had frequent responses to virion and immediate-early (IE) proteins of HSV-2, T cell responses in IS subjects were directed primarily at the early protein UL39 (ICP6) and the IE proteins ICP4 and ICP0. These data suggest that the antigenic repertoire in IS subjects is skewed compared to HSV-2+ subjects and thus, in Specific Aim 1, we will screen PBMC from IS and HLA-matched HSV-2+ control subjects using an HSV-2 ORFeome containing all predicted proteins of HSV-2. We will test the hypothesis that HSV-specific T cells from IS subjects are predominantly directed at IE proteins while HSV-specific T cells from HSV-2+ subjects are broadly directed at IE and virion proteins. The presence of frequent and persistent systemic T cell responses to HSV-2 in IS subjects suggests that T cells were primed in lymph nodes draining the genital mucosa after exposure to HSV-2 and in order for T cells to be protective, they must be present at local sites of antigen load, that is, in the genital mucosa and/or in genital skin. Although ES subjects lack detectable systemic T cell responses to HSV-2, it is possible that local T cells are present and thus, in Specific Aim 2, we will determine if HSV-specific T cells are present at the genital mucosa and in genital skin of IS and ES subjects. Defining the immunodominant HSV-2 proteins in IS subjects and the characterization of HSV-2 specific T cells at the genital mucosa in IS subjects would have a significant impact on the field of HSV-2 vaccine design where current vaccine strategies are modeled on T cell responses measured in HSV-2- infected subjects as opposed to T cell responses measured in subjects who appear to resist HSV infection.